期刊
JOURNAL OF BIOLOGICAL CHEMISTRY
卷 286, 期 8, 页码 6367-6374出版社
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M110.206227
关键词
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资金
- National Institutes of Health [R21HL087895, R01GM095538]
- Texas Norman Hackerman Advanced Research Program [010674-0034-2009]
The MelB permease of Salmonella typhimurium (MelB-ST) catalyzes the coupled symport of melibiose and Na+, Li+, or H+. In right-side-out membrane vesicles, melibiose efflux is inhibited by an inwardly directed gradient of Na+ or Li+ and stimulated by equimolar concentrations of internal and external Na+ or Li+. Melibiose exchange is faster than efflux in the presence of H+ or Na+ and stimulated by an inwardly directed Na+ gradient. Thus, sugar is released from MelB-ST externally prior to the release of cation in agreement with current models proposed for MelB of Escherichia coli (MelB-EC) and LacY. Although Li+ stimulates efflux, and an outwardly directed Li+ gradient increases exchange, it is striking that internal and external Li+ with no gradient inhibits exchange. Furthermore, Trp -> dansyl FRET measurements with a fluorescent sugar (2'-(N-dansyl) aminoalkyl-1-thio-beta-D-galactopyranoside) demonstrate that MelB-ST, in the presence of Na+ or Li+, exhibits K-app(d) values of similar to 1 mM for melibiose. Na+ and Li+ compete for a common binding pocket with activation constants for FRET of similar to 1 mM, whereas Rb+ or Cs+ exhibits little or no effect. Taken together, the findings indicate that MelB-ST utilizes H+ in addition to Na+ and Li+. FRET studies also show symmetrical emission maximum at similar to 500 nm with MelB-ST in the presence of 2'-(N-dansyl) aminoalkyl-1-thio-beta-D-galactopyranoside and Na+, Li+, or H+, which implies a relatively homogeneous distribution of conformers of MelB-ST ternary complexes in the membrane.
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